Since the normal organization and morphogenesis of epithelial tissue are under some control of a factor originating in mesenchymal cells, and diseases attributable to the morphogenetic abnormality of the epithelial tissue may be often caused by the mesenchymal cells present around the tissue, studies on the mechanism of mesenchymal cells which support the morphogenesis of the epithelial tissue have been made time. Although studies on the isolation, purification and structural analysis of a molecule, which controls the morphogenesis of the epithelial tissue, have been extensively made throughout the world, however, its substance has been scarcely known under the circumstances because the object of study is a substance which expresses under restrictions of time and space in a complicated system, and so it is difficult to produce a simplified experimental system.
In order to realize the elucidation of diseases caused by the morphogenetic abnormality of the epithelial tissue and the attack mechanism thereof, and the development of medical treatments for these diseases, it was an indispensable premise to isolate and purify such a molecule, which controls the morphogenesis of the epithelial tissue, and make the structure thereof clear. Therefore, an important problem in the art was to achieve the elucidation of the structure of such a molecule, and the like.
In such circumstances, the present inventors recently succeeded in isolation and identification of a molecule (the present inventors termed it "epimorphin") which controls the morphogenesis of the epithelial tissue (Japanese Patent Application Laid-Open No. 25295/1994). Epimorphin is a physiologically active substance comprising, as a core protein, a protein composed of 277 to 289 amino acids and is principally biosynthetically produced by mesenchymal cells.
The present inventors succeeded in determining the amino acid sequences of human and mouse epimorphin molecules. The splicing of their genes has revealed that at least three types exist in respective epimorphin molecules. The human epimorphin molecules include three types, human epimorphin represented by SEQ ID NO. 1, human epimorphin (isoform A) represented by SEQ ID NO. 2 and human epimorphin (isoform B) represented by SEQ ID NO. 3, all shown in SEQUENCE LISTING, which will be described subsequently. The mouse epimorphin molecules include three types, mouse epimorphin represented by SEQ ID NO. 4, mouse epimorphin (isoform A) represented by SEQ ID NO. 5 and mouse epimorphin (isoform B) represented by SEQ ID NO. 6, all shown in the SEQUENCE LISTING. The human epimorphin molecules and the mouse epimorphin molecules have homology of about 90% with each other at the amino acid level. Therefore, they are well conserved even between different animal species.
However, these epimorphin molecules involved a problem that since they firmly bind to a cell membrane at a domain (hereinafter referred to as "the C-terminal hydrophobic domain) adjacent to the C-terminus thereof, which is extremely high in hydrophobic nature, while taking a complex high-order structure in the living body so as to perform their functions, they are extremely difficult to prepare while keeping the activity of epimorphin at a high level. In particular, the epimorphin and the isoform A markedly show such a tendency. When a cell membrane-binding domain exists, it is difficult to secrete epimorphin produced by cultured animal cells into a medium so as to isolate and purify it. The present inventors proposed the preparation of a soluble modified epimorphin by a process of removing the C-terminal hydrophobic domain, and the like (Japanese Patent Application Laid-Open No. 25295/1994). However, these processes have been so far insufficient in the compatibility of the maintenance of high activity with the solubility, and there has hence been a demand for development of a more improved process.
If a modified epimorphin easy to prepare and purify can be obtained while keeping the physiological activity of epimorphin, it is useful in elucidating the attack mechanism of diseases caused by the morphogenetic abnormality of the epithelial tissue and developing medical treatments for these diseases.